[Objectives]This study was conducted to investigate the similarity and differences between TLR4 knockout mice and C57 BL/6 mice used in liver fibrosis research in terms of growth rate and reproduction ability.[Methods...[Objectives]This study was conducted to investigate the similarity and differences between TLR4 knockout mice and C57 BL/6 mice used in liver fibrosis research in terms of growth rate and reproduction ability.[Methods]Twenty TLR4 knockout mice and C57 BL/6 mice,half male and half female,were selected to compare the growth rates of body weight and body length of mice from the 4th to 12th weeks;and 20 pairs of male and female mice of the same strain were compared for the number of baby mice of the second litter.[Results]The growth rates of body weight and body length of the TLR4 knockout mice were significantly lower than those of C57 BL/6 mice(P<0.05)(except for the 4th and 5th weeks when there was no significant difference in body length);and in terms of reproductive ability,the TLR4 knockout mice were significantly lower than the C57 BL/6 mice(the ratio of the total number of baby mice in the second litter of the two strains,72∶147).[Conclusions]Knockout of the TLR4 gene has a significant impact on the growth and reproduction of mice.展开更多
Objective:To explore the protective effect and possible mechanism of action of Zhuyu Annao pill in mice with intracerebral hemorrhage(ICH).Methods:Sixty mice were divided into the control group,hemorrhage group,drug-t...Objective:To explore the protective effect and possible mechanism of action of Zhuyu Annao pill in mice with intracerebral hemorrhage(ICH).Methods:Sixty mice were divided into the control group,hemorrhage group,drug-treated group(after hemorrhage),TLR4-knockout hemorrhage group and TLR4-knockout hemorrhage + drug-treated group(after hemorrhage) with 12 in each group.Model of autologous ICH was established in all groups.After drilling and 12 h of fasting,models in the control group hemorrhage group and TLR4-knockout hemorrhage group were all drenched with 10 mL/kg distilled water by intragastric administration.Models in the drug-treated group and TLR4-knockout hemorrhage + drugtreated group were drenched with 6.25 g/kg of Zhuyu Annao pill.All groups were treated for 7 d.Longa scoring method was used to measure the neurological defect scores and determine the brain water contents of all groups;ELISA was employed to detect the inflammatory factor interleukin(IL)-6,tumor necrosis factor- α(TNF- α) and IL-1β in brain tissues;and Western blot was applied to test the expression quantities of apoptotic protein Bax and anti-apoptotic protein Bcl-2 in brain tissues.Results:At day 3 and7,compared with the hemorrhage group,the neurological defect scores of the drug-treated group,TLR4-knockout hemorrhage group and TLR4-knockout hemorrhage + drug-treated group decreased significantly(P<0.05) Compared with the hemorrhage group,the brain water contents of the drug-treated group,TLR4-knockout hemorrhage group and TLR4-knockout hemorrhage + drug-treated group reduced significantly(P<0.05) Compared with the hemorrhage group,the inflammatory factor IL-6,TNF-α and IL-1β of the drug-treated group,TLR4-knockout hemorrhage group and TLR4-knockout hemorrhage + drug-treated group decreased significantly(P<0.05).Compared with the hemorrhage group,the expression of apoptotic protein Bax of the drug-treated group,TLR4-knockout hemorrhage group and TLR4-knockout hemorrhage+ drug-treated group decreased significantly and the expression of anti-apoptotic protein Bcl-2 increased significantly(P<0.05).Conclusions:Zhuyu Annao pill can alleviateencephaledema for mice with ICH and reduce inflammatory responsesandnerve cell apoptosis.TLR4 can mediate inflammatory injury induced by ICH.Thus,Zhuyu Annao pill can play a protective role for brains by decreasing the expression of TLR4.展开更多
目的观察搜风祛痰中药复方稳斑汤对ApoE基因敲除小鼠动脉粥样硬化(AS)不稳定斑块动物模型的影响。方法 6～8周龄ApoE基因敲除小鼠制备AS不稳定斑块模型,分别给予阿托伐他汀以及不同剂量稳斑汤干预1个月,正常组采用生理盐水灌胃1个月。...目的观察搜风祛痰中药复方稳斑汤对ApoE基因敲除小鼠动脉粥样硬化(AS)不稳定斑块动物模型的影响。方法 6～8周龄ApoE基因敲除小鼠制备AS不稳定斑块模型,分别给予阿托伐他汀以及不同剂量稳斑汤干预1个月,正常组采用生理盐水灌胃1个月。取小鼠主动脉组织HE染色进行病理学观察,并采用免疫组化和RT-PCR技术检测HO-1和TLR4的表达水平。结果各组小鼠腹主动脉组织中HO-1的表达水平治疗组明显高于对照组(P<0.01),各组小鼠腹主动脉组织中TLR4的表达水平治疗组明显低于对照组(P<0.01)。结论 Apo E基因敲除小鼠AS不稳定斑块的形成与炎症因子密切相关,搜风祛痰中药复方稳斑汤可能通过调节HO-1和TLR4的表达而干预AS斑块形成及破裂。展开更多
[目的]构建敲除Toll Like Receptor 4(TLR4)基因的Ha Ca T细胞株,为后续利用该细胞株进行过敏原性研究提供材料。[方法]利用CRISPR/Cas9系统,根据靶向原理设计并合成4条特异性识别TLR4基因的向导RNA(single-molecule guide RNAs,sgRNAs)...[目的]构建敲除Toll Like Receptor 4(TLR4)基因的Ha Ca T细胞株,为后续利用该细胞株进行过敏原性研究提供材料。[方法]利用CRISPR/Cas9系统,根据靶向原理设计并合成4条特异性识别TLR4基因的向导RNA(single-molecule guide RNAs,sgRNAs),构建p X459-sgRNAh TLR4重组质粒,并转入Ha Ca T中,用嘌呤霉素筛选出单克隆阳性细胞。测序确认突变位点,然后利用脂多糖(lipopolysaccharide,LPS)刺激对TLR4进行功能验证来进一步确认敲除效果。[结果]测序结果表明#26单克隆细胞株在靶点附近缺失1 bp,造成TLR4编码基因的移码突变,蛋白翻译提前终止。功能性验证结果表明,在LPS的刺激下,IL-8和CCL20的mRNA水平分别下降约85%和90%,且IL-8的蛋白分泌水平也显著性下调(87%)。[结论]成功构建了敲除TLR4的稳定细胞株,并且验证TLR4的功能缺损。展开更多
基金National Natural Science Foundation of China(81960761,81960751,81902764)。
文摘[Objectives]This study was conducted to investigate the similarity and differences between TLR4 knockout mice and C57 BL/6 mice used in liver fibrosis research in terms of growth rate and reproduction ability.[Methods]Twenty TLR4 knockout mice and C57 BL/6 mice,half male and half female,were selected to compare the growth rates of body weight and body length of mice from the 4th to 12th weeks;and 20 pairs of male and female mice of the same strain were compared for the number of baby mice of the second litter.[Results]The growth rates of body weight and body length of the TLR4 knockout mice were significantly lower than those of C57 BL/6 mice(P<0.05)(except for the 4th and 5th weeks when there was no significant difference in body length);and in terms of reproductive ability,the TLR4 knockout mice were significantly lower than the C57 BL/6 mice(the ratio of the total number of baby mice in the second litter of the two strains,72∶147).[Conclusions]Knockout of the TLR4 gene has a significant impact on the growth and reproduction of mice.
基金supported by the system issue of Key Laboratory of Guangxi Basic Chinese Medicine Research(Grant No.16-380-58-06)
文摘Objective:To explore the protective effect and possible mechanism of action of Zhuyu Annao pill in mice with intracerebral hemorrhage(ICH).Methods:Sixty mice were divided into the control group,hemorrhage group,drug-treated group(after hemorrhage),TLR4-knockout hemorrhage group and TLR4-knockout hemorrhage + drug-treated group(after hemorrhage) with 12 in each group.Model of autologous ICH was established in all groups.After drilling and 12 h of fasting,models in the control group hemorrhage group and TLR4-knockout hemorrhage group were all drenched with 10 mL/kg distilled water by intragastric administration.Models in the drug-treated group and TLR4-knockout hemorrhage + drugtreated group were drenched with 6.25 g/kg of Zhuyu Annao pill.All groups were treated for 7 d.Longa scoring method was used to measure the neurological defect scores and determine the brain water contents of all groups;ELISA was employed to detect the inflammatory factor interleukin(IL)-6,tumor necrosis factor- α(TNF- α) and IL-1β in brain tissues;and Western blot was applied to test the expression quantities of apoptotic protein Bax and anti-apoptotic protein Bcl-2 in brain tissues.Results:At day 3 and7,compared with the hemorrhage group,the neurological defect scores of the drug-treated group,TLR4-knockout hemorrhage group and TLR4-knockout hemorrhage + drug-treated group decreased significantly(P<0.05) Compared with the hemorrhage group,the brain water contents of the drug-treated group,TLR4-knockout hemorrhage group and TLR4-knockout hemorrhage + drug-treated group reduced significantly(P<0.05) Compared with the hemorrhage group,the inflammatory factor IL-6,TNF-α and IL-1β of the drug-treated group,TLR4-knockout hemorrhage group and TLR4-knockout hemorrhage + drug-treated group decreased significantly(P<0.05).Compared with the hemorrhage group,the expression of apoptotic protein Bax of the drug-treated group,TLR4-knockout hemorrhage group and TLR4-knockout hemorrhage+ drug-treated group decreased significantly and the expression of anti-apoptotic protein Bcl-2 increased significantly(P<0.05).Conclusions:Zhuyu Annao pill can alleviateencephaledema for mice with ICH and reduce inflammatory responsesandnerve cell apoptosis.TLR4 can mediate inflammatory injury induced by ICH.Thus,Zhuyu Annao pill can play a protective role for brains by decreasing the expression of TLR4.
文摘目的观察搜风祛痰中药复方稳斑汤对ApoE基因敲除小鼠动脉粥样硬化(AS)不稳定斑块动物模型的影响。方法 6～8周龄ApoE基因敲除小鼠制备AS不稳定斑块模型,分别给予阿托伐他汀以及不同剂量稳斑汤干预1个月,正常组采用生理盐水灌胃1个月。取小鼠主动脉组织HE染色进行病理学观察,并采用免疫组化和RT-PCR技术检测HO-1和TLR4的表达水平。结果各组小鼠腹主动脉组织中HO-1的表达水平治疗组明显高于对照组(P<0.01),各组小鼠腹主动脉组织中TLR4的表达水平治疗组明显低于对照组(P<0.01)。结论 Apo E基因敲除小鼠AS不稳定斑块的形成与炎症因子密切相关,搜风祛痰中药复方稳斑汤可能通过调节HO-1和TLR4的表达而干预AS斑块形成及破裂。
文摘[目的]构建敲除Toll Like Receptor 4(TLR4)基因的Ha Ca T细胞株,为后续利用该细胞株进行过敏原性研究提供材料。[方法]利用CRISPR/Cas9系统,根据靶向原理设计并合成4条特异性识别TLR4基因的向导RNA(single-molecule guide RNAs,sgRNAs),构建p X459-sgRNAh TLR4重组质粒,并转入Ha Ca T中,用嘌呤霉素筛选出单克隆阳性细胞。测序确认突变位点,然后利用脂多糖(lipopolysaccharide,LPS)刺激对TLR4进行功能验证来进一步确认敲除效果。[结果]测序结果表明#26单克隆细胞株在靶点附近缺失1 bp,造成TLR4编码基因的移码突变,蛋白翻译提前终止。功能性验证结果表明,在LPS的刺激下,IL-8和CCL20的mRNA水平分别下降约85%和90%,且IL-8的蛋白分泌水平也显著性下调(87%)。[结论]成功构建了敲除TLR4的稳定细胞株,并且验证TLR4的功能缺损。
